The term “biomass” generally refers to the total amount of organic material that inhabits or exists in a given area. When such term is used for plants, in particular, it refers to dry weight per unit area. A biomass unit is quantified in terms of a mass or an energy amount. In the case of plant biomass, the term “standing crop” is occasionally used to represent “biomass.” Since plant biomass is generated by fixing atmospheric carbon dioxide with the use of solar energy, it can be regarded as so-called “carbon-neutral energy.” Accordingly, an increase in the amount of plant biomass is effective for global environmental protection, the prevention of global warming, and mitigation of greenhouse gas emissions. Thus, a technique for increasing the production of plant biomass is industrially significant.
Plants are cultivated for the purpose of using some tissue thereof (e.g., seeds, roots, leaves, or stems) or for the purpose of producing various materials, such as fat-and-oils. Examples of fat-and-oils produced from plants that have been heretofore known include soybean oil, sesame oil, olive oil, coconut oil, rice oil, cottonseed oil, sunflower oil, corn oil, safflower oil, palm oil, and rapeseed oil. Such fat-and-oils are extensively used for household and industrial applications. Also, fat-and-oils produced from plants are used as starting materials for biodiesel fuels or bioplastics, and the applicability thereof is increasing for alternative energy to petroleum.
Under such circumstances, it is necessary for the industrial success of the production of fat-and-oils using plants that the amount of production per unit of cultivation area be improved. If the number of cultivated plants is assumed to be constant per unit of cultivation area, an improvement in the amount of fat-and-oil production per plant is found to be necessary. When fat-and-oils are extracted from seeds obtained from plants, an improvement in the amount of fat-and-oil production per plant can be achieved via techniques of, for example, improving the seed yield per plant or increasing the fat-and-oil content in seeds.
Techniques for increasing the amount of fat-and-oil production from plant seeds are broadly classified into techniques based on an improvement in cultivation methods and techniques based on the development of plant varieties that can increase the amount of fat-and-oil production. Techniques based on the development of plant varieties that can increase the amount of fat-and-oil production are roughly classified as conventional breeding techniques such as crossing and molecular breeding techniques involving gene recombination. As techniques for increasing the amount of fat-and-oil production via gene recombination, A) a method of modifying a system for synthesizing triacylglycerol (TAG) of seeds, which is a main component of plant fat-and-oils, and B) a method of modifying a regulator gene that regulates plant morphogenesis or metabolism and expression of genes associated therewith are known.
In method A) above, the amount of TAG synthesized from a sugar produced via photosynthesis can be increased by (1) enhancing synthesis activity from a fatty acid or glycerol (i.e., TAG components) from sugars or (2) reinforcing the reaction of synthesizing TAG from glycerol and a fatty acid. In this regard, the following techniques have been reported as techniques that use genetic engineering techniques. An example of (1) is a technique in which cytosolic acetyl-coenzyme A carboxylase (ACCase) of Arabidopsis thaliana is overexpressed in rapeseed plastids and the fat-and-oil content in seeds is improved by 5% (Plant Physiology, 1997, Vol. 11, pp. 75-81). An example of (2) is a technique of increasing fat-and-oil production via overexpression of diacylglycerol acyltransferase (DGAT), which transfers an acyl group to the sn-3 position of diacylglycerol (Plant Physiology, 2001, Vol. 126, pp. 861-874). It is reported that the fat-and-oil content and the seed weight are increased as the DGAT expression level increases, and the number of seeds per plant may be occasionally increased according to the method of Plant Physiology, 2001, Vol. 126, pp. 861-874. The fat-and-oil content in Arabidopsis thaliana seeds was increased by 46% and the fat-and-oil amount per plant was increased by a maximum of about 125% by such technique.
As a form of method B), expression of transcriptional factor genes associated with the regulation of biosynthetic enzyme gene expression may be regulated. An example thereof can be found in WO 01/36597. WO 01/36597 employs a technique in which recombinant plants are prepared via exhaustive overexpression or knocking out of transcriptional factors and genes that enhance the fat-and-oil content in seeds are then selected. WO 01/36597 discloses that overexpression of ERF subfamily B-4 transcriptional factor genes results in a 23% increase in the fat-and-oil content in seeds. WO 01/36597, however, does not disclose an increase or decrease in fat-and-oil content per plant. Also, Plant J., 2004, 40, 575-585 discloses that the overexpression of WRINKLED1, which is a transcriptional factor having the AP2/EREB domain, improves the fat-and-oil content in seeds.
Although molecular breeding techniques as described above intended for the improvement of various traits have been developed, techniques for improving the yield involving increasing plant weight, increasing a given tissue, or improving the amount of target substances produced have not yet been put to practical use.
This is considered to be due to the following reasons. That is, truly adequate genes have not yet been discovered, and new recombinant varieties that are found effective at the test phase are unable to exhibit expected effects during the practical phase under a variety of natural environments. Also, many genes are associated with quantitative traits, such as increased plant weight, increased weight of a given tissue, or the amount of target substances produced, in various steps from those involving the control system to those involving the metabolic system, and it was difficult to discover and develop a truly adequate and useful gene that improves quantitative traits. In order to overcome such problems, the discovery of dramatically effective new genes and the development of genes exhibiting effects under practical environments, even if the effectiveness thereof is equivalent to that of existing genes, are necessary.